The mitochondrion is the center stage for energy metabolism, apoptosis, signaling, and ion homeostasis. Much of what we know about this organelle comes from studying mitochondrial respiratory chain disease (RCD). This devastating disease is due to genetic defects in the mtDNA or the nuclear DNA that give rise to a malfunctioning mitochondrial respiratory chain. Virtually all organ systems can be affected. RCD affects an estimated 1:5000 live births and is devastating - it is extremely difficult to diagnose, requiring consultation by multiple physicians and invasive biopsies, and at present no effective therapies are available. A small fraction of these disorders are maternally, but the vast majority of these disorders are due to mutations in nuclear genes, many of which have yet to be identified. Our research team has recently used integrative proteomics to define the ~1100 nuclear genes that encode the mitochondrial proteome - these genes represent a near-comprehensive collection of candidate genes for RCD. In the current application we have assembled a team consisting of leaders in mitochondrial medicine, computational genomics, and large-scale sequencing, to comprehensively resequence all these ~1100 nuclear genes in a panel of ~120 patients with clinically characterized RCD. Through this project, we solve the molecular bases for RCD, establish a facile, comprehensive DNA diagnostic test for RCD;and identify scores of new mitochondrial disease genes that will unlock new pockets of mitochondrial biology. PUBLIC HEALTH RELEVANCE: Mitochondrial disorders comprise one of the largest classes of inherited human disease, affecting both children and adults. We will analyze the DNA of patients with such disorders to discover their molecular basis. The results of this study may help us better diagnose and treat these devastating diseases.